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- EMDB-24177: Low conductance mechanosensitive channel YnaI -

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Basic information

Entry
Database: EMDB / ID: EMD-24177
TitleLow conductance mechanosensitive channel YnaI
Map data
Sample
  • Cell: Low conductance mechanosensitive channel YnaI
    • Protein or peptide: Low conductance mechanosensitive channel YnaI
  • Ligand: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine
Function / homology
Function and homology information


mechanosensitive monoatomic ion channel activity / cellular response to osmotic stress / identical protein binding / plasma membrane
Similarity search - Function
Mechanosensitive ion channel protein YnaI-like / : / : / Mechanosensitive ion channel MscS, C-terminal / Mechanosensitive ion channel, transmembrane helices 2/3 / Mechanosensitive ion channel MscS, conserved site / Uncharacterized protein family UPF0003 signature. / Mechanosensitive ion channel MscS, C-terminal / Mechanosensitive ion channel MscS, transmembrane-2 / Mechanosensitive ion channel MscS ...Mechanosensitive ion channel protein YnaI-like / : / : / Mechanosensitive ion channel MscS, C-terminal / Mechanosensitive ion channel, transmembrane helices 2/3 / Mechanosensitive ion channel MscS, conserved site / Uncharacterized protein family UPF0003 signature. / Mechanosensitive ion channel MscS, C-terminal / Mechanosensitive ion channel MscS, transmembrane-2 / Mechanosensitive ion channel MscS / Mechanosensitive ion channel, beta-domain / Mechanosensitive ion channel MscS, beta-domain superfamily / LSM domain superfamily
Similarity search - Domain/homology
Low conductance mechanosensitive channel YnaI
Similarity search - Component
Biological speciesEscherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Methodsingle particle reconstruction / cryo EM / negative staining / Resolution: 2.4 Å
AuthorsCatalano C / Ben-Hail D / Qiu W / des Georges A / Guo Y
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1R01GM132329 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1R35GM133598 United States
CitationJournal: Membranes (Basel) / Year: 2021
Title: Cryo-EM Structure of Mechanosensitive Channel YnaI Using SMA2000: Challenges and Opportunities.
Authors: Claudio Catalano / Danya Ben-Hail / Weihua Qiu / Paul Blount / Amedee des Georges / Youzhong Guo /
Abstract: Mechanosensitive channels respond to mechanical forces exerted on the cell membrane and play vital roles in regulating the chemical equilibrium within cells and their environment. High-resolution ...Mechanosensitive channels respond to mechanical forces exerted on the cell membrane and play vital roles in regulating the chemical equilibrium within cells and their environment. High-resolution structural information is required to understand the gating mechanisms of mechanosensitive channels. Protein-lipid interactions are essential for the structural and functional integrity of mechanosensitive channels, but detergents cannot maintain the crucial native lipid environment for purified mechanosensitive channels. Recently, detergent-free systems have emerged as alternatives for membrane protein structural biology. This report shows that while membrane-active polymer, SMA2000, could retain some native cell membrane lipids on the transmembrane domain of the mechanosensitive-like YnaI channel, the complete structure of the transmembrane domain of YnaI was not resolved. This reveals a significant limitation of SMA2000 or similar membrane-active copolymers. This limitation may come from the heterogeneity of the polymers and nonspecific interactions between the polymers and the relatively large hydrophobic pockets within the transmembrane domain of YnaI. However, this limitation offers development opportunities for detergent-free technology for challenging membrane proteins.
History
DepositionJun 4, 2021-
Header (metadata) releaseApr 20, 2022-
Map releaseApr 20, 2022-
UpdateApr 20, 2022-
Current statusApr 20, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_24177.png

Downloads & links

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Map

FileDownload / File: emd_24177.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy AUTHOR: 0.006
Minimum - Maximum-0.026699543 - 0.08194447
Average (Standard dev.)-8.625844e-05 (±0.0022871043)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 273.92 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_24177_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_24177_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Low conductance mechanosensitive channel YnaI

EntireName: Low conductance mechanosensitive channel YnaI
Components
  • Cell: Low conductance mechanosensitive channel YnaI
    • Protein or peptide: Low conductance mechanosensitive channel YnaI
  • Ligand: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine

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Supramolecule #1: Low conductance mechanosensitive channel YnaI

SupramoleculeName: Low conductance mechanosensitive channel YnaI / type: cell / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)

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Macromolecule #1: Low conductance mechanosensitive channel YnaI

MacromoleculeName: Low conductance mechanosensitive channel YnaI / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Strain: K12
Molecular weightTheoretical: 38.792344 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MIAELFTNNA LNLVIIFGSC AALILMSFWF RRGNRKRKGF LFHAVQFLIY TIIISAVGSI INYVIENYKL KFITPGVIDF ICTSLIAVI LTIKLFLLIN QFEKQQIKKG RDITSARIMS RIIKITIIVV LVLLYGEHFG MSLSGLLTFG GIGGLAVGMA G KDILSNFF ...String:
MIAELFTNNA LNLVIIFGSC AALILMSFWF RRGNRKRKGF LFHAVQFLIY TIIISAVGSI INYVIENYKL KFITPGVIDF ICTSLIAVI LTIKLFLLIN QFEKQQIKKG RDITSARIMS RIIKITIIVV LVLLYGEHFG MSLSGLLTFG GIGGLAVGMA G KDILSNFF SGIMLYFDRP FSIGDWIRSP DRNIEGTVAE IGWRITKITT FDNRPLYVPN SLFSSISVEN PGRMTNRRIT TT IGLRYED AAKVGVIVEA VREMLKNHPA IDQRQTLLVY FNQFADSSLN IMVYCFTKTT VWAEWLAAQQ DVYLKIIDIV QSH GADFAF PSQTLYMDNI TPPEQGR

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Macromolecule #2: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine

MacromoleculeName: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine / type: ligand / ID: 2 / Number of copies: 7 / Formula: PEE
Molecular weightTheoretical: 744.034 Da
Chemical component information

ChemComp-PEE:
1,2-dioleoyl-sn-glycero-3-phosphoethanolamine / DOPE, phospholipid*YM / Discrete optimized protein energy

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.8
StainingType: NEGATIVE / Material: 2 % uranil acetate
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 2.0 nm / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 2155 / Average exposure time: 8.0 sec. / Average electron dose: 56.18 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 330000
CTF correctionSoftware - Name: CTFFIND (ver. 4.1)
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final reconstructionApplied symmetry - Point group: C7 (7 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 142000

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Atomic model buiding 1

Initial modelPDB ID:

5y4o

RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-7n4t:
Low conductance mechanosensitive channel YnaI

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